title: The hexosamine biosynthetic pathway can mediate myocardial apoptosis in a rat model of diet-induced insulin resistance.
abstract: OBJECTIVE
  Type 2 diabetes is characterized by deranged metabolic pathways that may result in cardiovascular complications. For example, hyperglycaemia promotes flux through the hexosamine biosynthetic pathway (HBP) leading to greater O-GlcNAcylation of target proteins, with pathophysiological outcomes. This study investigated mechanisms whereby increased HBP flux elicits myocardial apoptosis in a rat model of diet-induced hyperglycaemia/insulin resistance.
  
  
  METHODS
  Four-week-old male Wistar rats were fed a high-fat diet (86 days) after which insulin resistance was assessed vs. matched controls. Oxidative stress was evaluated, and apoptotic peptide levels, BAD phosphorylation and overall O-GlcNAcylation assessed by immunoblotting. Protein-specific O-GlcNAcylation and BAD-Bcl-2 dimerization were determined by immunoprecipitation and Western blotting.
  
  
  RESULTS
  Rats consuming the high-fat diet exhibited a moderate elevation in body weight, higher fasting insulin and glucose levels, and insulin resistance vs. controls. Overall protein O-GlcNAcylation was increased in hyperglycaemic/insulin-resistant hearts. In parallel, myocardial peptide levels of apoptotic markers (caspase-3, cytochrome-c, BAD) were significantly higher with insulin resistance. To gain mechanistic insight into our findings, we evaluated O-GlcNAcylation of BAD, a pro-apoptotic Bcl-2 homolog. Here we found increased BAD O-GlcNAcylation and decreased BAD phosphorylation (Ser136) in hyperglycaemic/insulin-resistant rat hearts. These data are in agreement with competition by phosphorylation and O-GlcNAcylation for the same or neighbouring site(s) on target proteins. Moreover, we observed increased BAD-Bcl-2 dimerization in hyperglycaemic/insulin-resistant hearts.
  
  
  CONCLUSIONS
  The main finding of this study is that increased apoptosis in hyperglycaemic/insulin-resistant hearts can also be mediated through HBP-induced BAD O-GlcNAcylation and greater formation of BAD-Bcl-2 dimers (pro-apoptotic).
authors.affiliation: Cardio-Metabolic Research Group, Department of Physiological Sciences, Stellenbosch University, South Africa.
authors.forename: U
authors.initials: U
authors.lastname: Rajamani
authors.forename: D
authors.initials: D
authors.lastname: Joseph
authors.forename: S
authors.initials: S
authors.lastname: Roux
authors.forename: M F
authors.initials: MF
authors.lastname: Essop
mesh_terms: D000818:Animals; D017209:Apoptosis / Q000502:physiology*; D015415:Biomarkers / Q000378:metabolism; D053898:Biosynthetic Pathways / Q000502:physiology; D003924:Diabetes Mellitus, Type 2 / Q000378:metabolism; D004032:Diet / Q000009:adverse effects*; D004041:Dietary Fats; D006595:Hexosamines / Q000096:biosynthesis*; D006801:Humans; D006943:Hyperglycemia / Q000378:metabolism; D007333:Insulin Resistance / Q000502:physiology*; D008297:Male; D009206:Myocardium / Q000378:metabolism*; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000378:metabolism; D051381:Rats; D017208:Rats, Wistar; D051025:bcl-Associated Death Protein / Q000378:metabolism
chemical_list: D015415:Biomarkers; D004041:Dietary Fats; D006595:Hexosamines; D019253:Proto-Oncogene Proteins c-bcl-2; D051025:bcl-Associated Death Protein